We (R.L. Ax and R.J. Ryan) recently completed a study of the role of FSH on proteoglycan synthesis in vivo and in vitro. Briefly, the concentration of the mucopolysaccharides (carbohydrate portions of proteoglycans) chondroitin sulfate and heparan sulfate paralled the reported numbers of FSH receptors and FSH-stimulatable adenylate cyclase in granulosa cells. The degree of sulfation of the chondroitin increased with follicular maturation. In vitro studies showed FSH stimulated 3H-glucosamine incorporation into proteoglycans in a linear log-dose response. The response was not elicited by LH, HCG or TSH. Furthermore, dibutyrl cyclic AMP mimicked the FSH action. Inhibitors of protein synthesis or RNA synthesis decreased the response when administereed within the first 8 hrs, after FSH. If this grant proposal is approved, studies would continue in two major areas of effort. Dr. R. J. Ryan at Mayo Clinic would initiate exeriments: (1) to purify the proteoglycan(s) from follicular fluid. Once purified they would be (2) chemically characterized, (3) used as antigens plus or minus carbohydrate moiety to make antibodies, and antibodies would be used to (4) precipitate radio-labelled proteoglycans from in vitro studies. Dr. R. L. Ax at the University of Wisconsin would: (1) validate the current in vitro system for use as a routine bioassay for FSH, (2) try various labelled carbohydrates and amino acids to calculate their efficiency of incorporation to deduce rate-limiting step(s) of the site of FSH action, (3) chemically characterize fluid from cystic and atretic follicles of farm animals in comparison with fluid from normal follicles to see what changes occur to proteoglycans in ovarian dysfunction and (4) investigate proteolgycan importance of proteoglycans to follicular fate, and compare proteoglycans of normal follicles to abnormal follicles.